The 26S proteasome performs regulated proteolysis in eukaryotes and has emerged as a major therapeutic target. It is composed of a 20S catalytic core particle (CP) where protein degradation occurs, that is capped at either end by a 19S regulatory particle. Proteasome substrates are distinguished by post-translational modification with ubiquitin chains, which bind directly to receptor sites in the RP. A major goal of our section is to understand how the proteasome recognizes substrates and becomes committed to substrate proteolysis.
We aim to use this knowledge to design new methods that target specific components of the ubiquitin-proteasome pathway. In collaboration with extramural researchers, we established Rpn1 and Rpn13 as substrate receptors in the proteasome that can directly interact with ubiquitin or ubiquitin-like domains of shuttle factors that contain ubiquitin-associated domains. We used NMR spectroscopy to solve the structure of the three major substrate receptor sites in the RP (Rpn1, Rpn10 and Rpn13) complexed with ubiquitin chains. This work is being extended to evaluate the specific functional roles of Rpn1, Rpn10 and Rpn13, both by biophysical methods and cell biology techniques. In addition, we are further defining proteasome structure at atomic level resolution. In a related collaborative project, we found that a class of bis-benzylidine piperidone derivatives trigger effects similar to proteasome inhibition, including restriction of tumor growth in mouse xenograft models. In contrast to the FDA-approved inhibitors, this class of small molecules (exemplified by RA190) do not inhibit the proteasome CP and instead covalently attach to hRpn13, which we have found is required for RA190 sensitivity. We are actively pursuing the mechanistic relationship between RA190 and hRpn13. Altogether, this project is providing fundamental information on how proteasome recognizes ubiquitinated substrates at atomic level resolution, as well as the therapeutic potential of targeting alternative sites in the proteasome.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIABC011490-06
Application #
9779931
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
6
Fiscal Year
2018
Total Cost
Indirect Cost
Name
Basic Sciences
Department
Type
DUNS #
City
State
Country
Zip Code
Calabrese, David R; Chen, Xiang; Leon, Elena C et al. (2018) Chemical and structural studies provide a mechanistic basis for recognition of the MYC G-quadruplex. Nat Commun 9:4229
Lu, Xiuxiu; Nowicka, Urszula; Sridharan, Vinidhra et al. (2017) Structure of the Rpn13-Rpn2 complex provides insights for Rpn13 and Uch37 as anticancer targets. Nat Commun 8:15540
Chen, Xiang; Randles, Leah; Shi, Ke et al. (2016) Structures of Rpn1 T1:Rad23 and hRpn13:hPLIC2 Reveal Distinct Binding Mechanisms between Substrate Receptors and Shuttle Factors of the Proteasome. Structure 24:1257-1270
Rao, Timsi; Gao, Rui; Takada, Saeko et al. (2016) Novel TDP2-ubiquitin interactions and their importance for the repair of topoisomerase II-mediated DNA damage. Nucleic Acids Res 44:10201-10215
Finley, Daniel; Chen, Xiang; Walters, Kylie J (2016) Gates, Channels, and Switches: Elements of the Proteasome Machine. Trends Biochem Sci 41:77-93
Randles, Leah; Anchoori, Ravi K; Roden, Richard B S et al. (2016) The Proteasome Ubiquitin Receptor hRpn13 and Its Interacting Deubiquitinating Enzyme Uch37 Are Required for Proper Cell Cycle Progression. J Biol Chem 291:8773-83
Shi, Yuan; Chen, Xiang; Elsasser, Suzanne et al. (2016) Rpn1 provides adjacent receptor sites for substrate binding and deubiquitination by the proteasome. Science 351:
Chen, Xiang; Walters, Kylie J (2016) (1)H, (15)N, (13)C resonance assignments for Saccharomyces cerevisiae Rad23 UBL domain. Biomol NMR Assign 10:291-5
Nowicka, Urszula; Hoffman, Morgan; Randles, Leah et al. (2016) Mycobacterium tuberculosis copper-regulated protein SocB is an intrinsically disordered protein that folds upon interaction with a synthetic phospholipid bilayer. Proteins 84:193-200
Yu, Clinton; Yang, Yingying; Wang, Xiaorong et al. (2016) Characterization of Dynamic UbR-Proteasome Subcomplexes by In vivo Cross-linking (X) Assisted Bimolecular Tandem Affinity Purification (XBAP) and Label-free Quantitation. Mol Cell Proteomics 15:2279-92

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